Nakhoul N L, Abdulnour-Nakhoul S, Khuri R N, Lieberman E M, Hargittai P T
Department of Medicine, East Carolina University School of Medicine, Greenville, North Carolina 27858-4354.
Glia. 1994 Mar;10(3):155-64. doi: 10.1002/glia.440100302.
We examined H+ and HCO3- transport mechanisms that are involved in the regulation of intracellular pH of Schwann cells. Primary cultures of Schwann cells were prepared from the sciatic nerves of 1-3-day-old rats. pHi of single cells attached to cover slips was continuously monitored by measuring the absorbance spectra of the pH-sensitive dye dimethylcarboxyfluorescein incorporated intracellularly. The average pHi of neonatal Schwann cells bathed in HEPES mammalian solution was 7.17 +/- 0.02 (n = 32). In the nominal absence of HCO3-, pHi spontaneously recovered from an acute acid load induced by exposing the Schwann cells to 20 mM NH4+ (NH4+ prepulse). This pHi recovery from the acute acid load was totally inhibited in the absence of external Na+ or in the presence of 1 mM amiloride. In both cases, the pHi recovery was readily restored upon readdition of external Na+ or removal of amiloride. In the steady-state, addition of amiloride caused a small and slow decrease in pHi which was readily reversed upon removal of amiloride. In the presence of HCO3-, removal of external Cl- caused pHi to rapidly and reversibly increase by 0.23 +/- 0.03 (n = 15) and the initial rate of alkalinization was 20.6 +/- 2.7 x 10(-4) pH/sec. In the absence of external Na+, removal of bath Cl- still caused pHi to increase by 0.15 +/- 0.02 and the initial rate of pHi increase was not significantly altered. In the nominal absence of HCO3-, removal of bath Cl- caused pHi to increase very slightly (0.05 +/- 0.01) with an initial dpHi/dt of only 4.4 +/- 0.2 x 10(-4) pH/sec (n = 4). Addition of 100 microM DIDS did not inhibit the pHi increase caused by removal of bath Cl-. These data indicate that 1) Rat Schwann cells regulate their pHi via an Na-H exchange mechanism which is moderately active at steady-state pHi. 2) In the presence of HCO3-, there is a Na-independent Cl-HCO3 (base) exchanger which also contributes to regulation of intracellular pH in Schwann cells.
我们研究了参与雪旺细胞胞内pH调节的H⁺和HCO₃⁻转运机制。雪旺细胞原代培养物取自1 - 3日龄大鼠的坐骨神经。通过测量细胞内掺入的pH敏感染料二甲基羧基荧光素的吸收光谱，连续监测附着在盖玻片上的单个细胞的pHi。在HEPES哺乳动物溶液中培养的新生雪旺细胞的平均pHi为7.17±0.02（n = 32）。在名义上不存在HCO₃⁻的情况下，将雪旺细胞暴露于20 mM NH₄⁺（NH₄⁺预脉冲）诱导的急性酸负荷后，pHi会自发恢复。在没有外部Na⁺或存在1 mM氨氯吡脒的情况下，急性酸负荷后的pHi恢复完全受到抑制。在这两种情况下，重新添加外部Na⁺或去除氨氯吡脒后，pHi恢复很容易恢复。在稳态下，添加氨氯吡脒会导致pHi缓慢小幅下降，去除氨氯吡脒后很容易逆转。在存在HCO₃⁻的情况下，去除外部Cl⁻会导致pHi迅速且可逆地升高0.23±0.03（n = 15），碱化的初始速率为20.6±2.7×10⁻⁴pH/秒。在没有外部Na⁺的情况下，去除浴液中的Cl⁻仍会导致pHi升高0.15±0.02，pHi升高的初始速率没有显著改变。在名义上不存在HCO₃⁻的情况下，去除浴液中的Cl⁻会导致pHi非常轻微地升高（0.05±0.01），初始dpHi/dt仅为4.4±0.2×10⁻⁴pH/秒（n = 4）。添加100μM DIDS不会抑制去除浴液中的Cl⁻引起的pHi升高。这些数据表明：1）大鼠雪旺细胞通过Na⁺ - H⁺交换机制调节其pHi，该机制在稳态pHi时具有适度活性。2）在存在HCO₃⁻的情况下，存在一种不依赖Na⁺的Cl⁻ - HCO₃⁻（碱）交换器，它也有助于雪旺细胞内pH的调节。
